The Ubiquitin-Proteasome System in Metazoan Embryogenesis Much of our understanding about the genetic regulation of development centers around gene regulatory networks that drive different transcriptional cascades with temporal and spatial specificity. Proteolysis through the ubiquitin-proteasome system (UPS) has emerged as an essential layer of regulation to remove specific signaling molecules and transcriptional regulators at specific stages of development to ensure proper progression of developmental events. Using C. elegans embryogenesis as a model, we will take a novel multidisciplinary approach to systematically and rapidly elucidate the in vivo functions of the UPS in Metazoan development. We have identified ~50 conserved, embryonic lethal genes that are required for UPS substrate specificity. In addition, we have developed novel technologies to automatically track every cell at every minute through embryogenesis, and will further develop methods to systematically assay the behaviors of every cell based on quantitative measurements of cell division, fate marker expression and cell movement. Using these technologies, our genetic and phenotypic analysis will define the specific developmental contexts and processes that these ~50 conserved genes regulate at single-cell resolution. We will then take a systems biology approach to construct a predicted interaction network of these genes with key developmental regulators and pathways based on phenotypic similarity, and integrate this developmental network with the large existing human gene interaction network based on sequence orthology. In doing so, we will build an extensive gene network that connects UPS genes and their interactors to specific developmental processes and their underlying regulatory pathways. This network will therefore be particularly useful in guiding future studies of how the UPS regulates human development and disease. Finally, we will conduct an in depth study on how the UPS controls the balance between self renewal and differentiation in the early C. elegans embryo, a novel and unexpected aspect of C. elegans development revealed in our pilot studies of the UPS. PUBLIC HEALTH RELEVANCE: The Ubiquitin Proteasome System is a complex regulatory network in cells. Using a Systems Biology strategy and a multidisciplinary approach involving live microscopy, genetics and computational sciences, we will link these key regulators to developmental processes and human pathology, such as over proliferation in cancer.